Project Summary/Abstract The long-term objective of this proposal is to understand induction and effector mechanisms of immunity in the absence of signaling adaptor MyD88. We use as a tool the intracellular protozoan Toxoplasma gondii, an orally acquired opportunistic pathogen that normally induces strong protective Th1- and IFN-?-based immunity. Toxoplasma can activate mouse immunity through an MyD88-dependent TLR11/12 pathway. Yet, humans lack these TLR and populations deficient in MyD88 expression retain resistance to all but a limited number of bacterial infections. Therefore, immune pathways that do not involve MyD88 are important to understand. We have found that MyD88 knockout mice are capable of generating Th1-like immunity during Toxoplasma infection, indicating that alternative pathways for immune initiation are active in the mouse model. In addition, MyD88 knockout animals can be vaccinated against lethal infection with orally inoculated T. gondii. The cell and molecular mechanisms underpinning MyD88-independent immunity are uncharacterized. The central hypothesis driving this research proposal is that the mouse immune system is equipped to generate protective immunity to Toxoplasma using MyD88-independent pathways of resistance. We more generally hypothesize that this is important for understanding human disease based upon overall resistance to infections in the absence of MyD88. Three Specific Aims will be used to understand MyD88-independent immunity during infection. Aim 1: Determine how T cells are activated in the absence of MyD88 signaling in the mucosal response to Toxoplasma. We will investigate cytokine requirements for MyD88-independent generation of IFN-?-positive T cells and characterize the phenotype and specificity of these cells. Aim 2: Determine the impact of MyD88 deficiency on gut microbicidal effector functions. We will examine how deficiency in this signaling adaptor affects recruitment and function of anti- microbial effector cells in the intestinal mucosa following Toxoplasma infection. Aim 3: Determine how vaccination with an avirulent Toxoplasma strain triggers MyD88-independent protective immunity in the intestinal mucosa. We will employ a nonproliferating Toxoplasma strain to determine how T. gondii induces resistance to lethal oral challenge in a model where immunity can be triggered without complications associated with parasite-induced tissue damage. The importance of this research is that it will significantly extend and deepen our understanding of induction and effector mechanisms of immunity in the absence of MyD88, which is critical to understanding host defense in humans. The ultimate impact of this research is that it can be expected to identify novel parasite and host targets for promoting resistance and immunity during infectious disease.